OBJECTIVES: a. We propose to carry out studies of the operation and control of the purine nucleotide cycle in perfused skeletal muscle. At rest our preparations produce little ammonia. Electrical stimulation results in a considerable output of ammonia. The objective is to understand the conditions under which the purine nucleotide cycle operates, and how its operation affects the control of glycolysis and the citric acid cycle. b. We propose to study how the interactions of the purine nucleotide cycle with glycolysis lead to oscillations in both pathways. The effect of variables such as pH and the degree of energy drain on the rates of operation of the purine nucleotide cycle and glycolysis, and on the type of oscillation, will be investigated in extracts prepared from different types of muscles, in different nutritional and hormonal states. Initial studies demonstrating the operation of the cycle in extracts of brain and kidney have been completed. The purine nucleotide cycle can operate at a rate adequate to account for the maximum rates of ammonia production by brain, for 100% of ammonia production by kidney of normal rats, and 64% of ammonia production by kidney of acidotic rats. The control properties of the cycle in brain extracts are being studied because they differ from those of muscle (adenylate deaminases from brain and muscle possess different regulatory properties.) c. The regulatory properties of adenylosuccinate synthetase and adenylate deaminase are being studied so that the factors that control the operation of the purine nucleotide cycle can be better understood. BIBLIOGRAPHIC REFERENCES: "Purine Nucleotide Cycle. Evidence for the Occurrence of the Cycle in Brain" by Vera Schultz and John M. Lowenstein (1976) J. Biol. Chem. 251, 485-492. "Insulin Removal by Isolated Perfused Rat Liver" by Robert I. Misbin, Thomas J. Merimee, and John M. Lowenstein (1976) Amer. J. Physiol. 230, 171-177.